Project Summary Food allergy is rapidly increasing in prevalence and is the most common cause of anaphylaxis. While significant progress has been made in our understanding of the underlying immunologic pathways involved in dendritic cell (DC) presentation of food allergens, the development of the CD4+ Th2 repertoire, and the IgE-MC?dependent effector processes, there has been little attention to the processes underlying the passage of food allergens across the GI epithelium and presentation of these allergens to the immune compartment. We have recently reported 1) that goblet cells (GCs) in the small intestine (SI) of nave mice can act as a conduit and permit the passage of food allergens across the intestinal epithelial (IE) layer and presentation to the SI lamina propria immune compartment (termed Goblet cell antigen passages; GAPs)13; 2) that food allergic mice utilize a different IE transport mechanism involving SI villus and crypt GCs, enteroendocrine cells, and Paneth cells that passage food allergens across the intestinal epithelial layer (termed secretory epithelial antigen passages; SAPs)24 and 3) a critical role for IE cell-derived pro-Type 2 cytokines such as TSLP, IL-25 and IL-33 in the development of food-induced anaphylaxis in mice14. In a series of preliminary studies, we interconnect these observations demonstrating that induction of SAPs and uptake of food allergens in murine intestinal epithelial cells leads to expression of the pro-allergic cytokine, IL- 33 and employing a human intestinal organoid (HIO) transplant model system demonstrate that the molecular processes, GAPs and SAPs are conserved in humans. The current gap in knowledge is the specificity of food allergen uptake by secretory intestinal epithelial cell lineages; the potential existence of environmental trigger programming of IEs which leads to modified antigen passage patterning (GAPs vs SAPs); and the relationship between food allergen uptake by SAPs and expression of pro-Type 2 cytokines in human tissue. We hypothesize that SAPs are a mechanism by which food allergens are channeled across the intestinal epithelium, promote the production of pro-Type 2 cytokines and whose composition and function are modulated by environmental triggers. In Aim I we will define the SI secretory cell lineages involved in food allergen passages and the impact of environmental triggers on food allergen passage patterning and Aim II, define the transcriptional inflammatory signature of human intestinal epithelial cells following food allergen uptake. With respect to the expected outcomes, the studies proposed in Aim I are expected to identify the involvement of distinct SI epithelial-specific food allergen transport processes and how they are influenced by environmental triggers, and those in Aim II are expected to reveal a link between pro-Th2 cytokine production and food allergen uptake by SI intestinal epithelial cells and define the pro-allergic transcriptional inflammatory signature of antigen passages. Collectively, these studies will illuminate new IE-restricted processes by which food allergens are sampled by the human SI compartment; link food allergen uptake with pro-Type 2 cytokine production and identify divergent pathways of SI IEs food allergen uptake that are influenced by FA status.